Fluorescence-Activated Cell Sorting–Based Purification of Embryonic Stem Cell–Derived Neural Precursors Averts Tumor Formation after Transplantation

Abstract

The differentiation of dopaminergic (DA) neurons from mouse embryonic stem cells (ESCs) can be efficiently induced, making these neurons a potential source for transplantation as a treatment for Parkinson's disease, a condition characterized by the gradual loss of midbrain DA neurons. One of the major persistent obstacles to the successful implementation of therapeutic ESC transplantation is the propensity of ESC-derived grafts to form tumors in vivo. To address this problem, we used fluorescence-activated cell sorting to purify mouse ESC-derived neural precursors expressing the neural precursor marker Sox1. ESCderived, Sox1 ؉ cells began to express neuronal cell markers and differentiated into DA neurons upon transplantation into mouse brains but did not generate tumors in this site. In contrast, Sox1 ؊ cells that expressed ESC markers frequently formed tumors in vivo. These results indicate that Sox1-based cell sorting of neural precursors prevents graftderived tumor formation after transplantation, providing a promising strategy for cell transplantation therapy of neurodegenerative disorders.

Transplantation of telencephalic neural progenitors induced from embryonic stem cells into subacute phase of focal cerebral ischemia

“…sigmaaldrich.com), were used in order to identify the transplanted cells, because after transplantation Sox1-GFP ES cells would fade GFP expression with their differentiation. 17 All recombinant DNA research conformed to National Institutes of Health (NIH) guidelines.…”

“…To separate two distinct cell populations, Sox1-GFP-positive and Sox1-GFP-negative cells were sorted using a FACSAria cell sorter and FACSDiva software (Beckton Dickinson, Franklin Lakes, NJ, USA) as described previously. 17 Dead ES cells were identified and eliminated by propidium iodide staining and forward-side scatter gating, respectively. Gates for each population were set so that the two subsets sorted based on Sox1 staining would not overlap when reanalyzed.…”

“…The observation of a Ki67-positive cell mass was defined as positive tumor formation. 17 Teratoma volume was evaluated in the same way as graft size.…”

“…We have previously confirmed that over 90% of the GFP-positive cells differentiated into Tuj1-positive postmitotic neurons, and half of the GFP-negative cells expressed the Oct4. 17 ES cell-derived telencephalic neural progenitors for cerebral ischemia M Fujimoto et al…”

“…17 However, the behavior of neural progenitors sorted by Sox1 in ischemic brain has not been clarified. This is the first report that shows SFEB culture could induce the striatum neurons from ES cells effectively both in vitro and in vivo and Sox1-based cell sorting of neural progenitors prevented tumor formation in ischemic brain for 8 weeks.…”